Adipose tissue is a critical regulator of osteoarthritis.

Osteoarthritis (OA), the leading cause of pain and disability worldwide, disproportionally affects individuals with obesity. The mechanisms by which obesity leads to the onset and progression of OA are unclear due to the complex interactions among the metabolic, biomechanical, and inflammatory factors that accompany increased adiposity. We used a murine preclinical model of lipodystrophy (LD) to examine the direct contribution of adipose tissue to OA. Knee joints of LD mice were protected from spontaneous or posttraumatic OA, on either a chow or high-fat diet, despite similar body weight and the presence of systemic inflammation. These findings indicate that adipose tissue itself plays a critical role in the pathophysiology of OA. Susceptibility to posttraumatic OA was reintroduced into LD mice using implantation of a small adipose tissue depot derived from wild-type animals or mouse embryonic fibroblasts that undergo spontaneous adipogenesis, implicating paracrine signaling from fat, rather than body weight, as a mediator of joint degeneration.

Role of Mineralocorticoid Receptor in Adipogenesis and Obesity in Male Mice.

Increased visceral adiposity and hyperglycemia, 2 characteristics of metabolic syndrome, are also present in conditions of excess glucocorticoids (GCs). GCs are hormones thought to act primarily via the glucocorticoid receptor (GR). GCs are commonly prescribed for inflammatory disorders, yet their use is limited due to many adverse metabolic side effects. In addition to GR, GCs also bind the mineralocorticoid receptor (MR), but there are many conflicting studies about the exact role of MR in metabolic disease. Using MR knockout mice (MRKO), we find that both white and brown adipose depots form normally when compared with wild-type mice at P5. We created mice with adipocyte-specific deletion of MR (FMRKO) to better understand the role of MR in metabolic dysfunction. Treatment of mice with excess GCs for 4 weeks, via corticosterone in drinking water, induced increased fat mass and glucose intolerance to similar levels in FMRKO and floxed control mice. Separately, when fed a high-fat diet for 16 weeks, FMRKO mice had reduced body weight, fat mass, and hepatic steatosis, relative to floxed control mice. Decreased adiposity likely resulted from increased energy expenditure since food intake was not different. RNA sequencing analysis revealed decreased enrichment of genes associated with adipogenesis in inguinal white adipose of FMRKO mice. Differentiation of mouse embryonic fibroblasts (MEFs) showed modestly impaired adipogenesis in MRKO MEFs compared with wild type, but this was rescued upon the addition of peroxisome proliferator-activated receptor gamma (PPARγ) agonist or PPARγ overexpression. Collectively, these studies provide further evidence supporting the potential value of MR as a therapeutic target for conditions associated with metabolic syndrome.

Epithelial-adipocyte interactions are required for mammary gland development, but not for milk production or fertility.

To investigate the role of adipose tissue in reproductive function and mammary gland development and function, we have examined lipodystrophic (LD) mice. LD mice of both sexes are sterile, but fertility can be restored with leptin injections. Mammary glands from lipodystrophic mice were rudimentary and lacked terminal end buds. Leptin-injected LD mice were able to become pregnant, showed normal pregnancy-associated glandular proliferation despite a smaller glandular area, were able to produce a small amount of milk that had grossly normal content of milk proteins and neutral lipids, but could not sustain pups to weaning. In order to separate the individual requirements for 1) adipokines such as leptin, 2) estradiol, and 3) physical epithelial-adipocyte interactions, we performed a series of experiments with both lipodystrophic mice and ob (obese mice with a mutation in the lep gene encoding the adipokine leptin) mice that received either estradiol treatment or preadipocyte transplant. The resulting fat pad did not rescue the defect in mammary gland development in lipodystrophic mice. The defect also could not be rescued with estradiol pellets. Ob/ob mice, like LD mice, lack leptin and estradiol, but retain adipose tissue. Ob mice have defective mammary gland development. However, in striking contrast to what was observed in lipodystrophic mice, reconstitution of a WT fat pad in ob mice rescued the defect in mammary gland development. Estradiol treatment did not rescue mammary gland development in ob mice. Therefore direct interaction between mammary gland epithelia and adipocytes is a requirement for full invasion and expansion of the gland, but is not required for glandular proliferation during pregnancy and milk production.

Mouse Embryonic Fibroblasts Protect ob/ob Mice From Obesity and Metabolic Complications.

The global obesity epidemic is fueling alarming rates of diabetes, associated with increased risk of cardiovascular disease and cancer. Leptin is a hormone secreted by adipose tissue that is a key regulator of body weight (BW) and energy expenditure. Leptin-deficient humans and mice are obese, diabetic, and infertile and have hepatic steatosis. Although leptin replacement therapy can alleviate the pathologies seen in leptin-deficient patients and mouse models, treatment is costly and requires daily injections. Because adipocytes are the source of leptin secretion, we investigated whether mouse embryonic fibroblasts (MEFs), capable of forming adipocytes, could be injected into ob/ob mice and prevent the metabolic phenotype seen in these leptin-deficient mice. We performed a single subcutaneous injection of MEFs into leptin-deficient ob/ob mice. The MEF injection formed a single fat pad that is histologically similar to white adipose tissue. The ob/ob mice receiving MEFs (obRs) had significantly lower BW compared with nontreated ob/ob mice, primarily because of decreased adipose tissue mass. Additionally, obR mice had significantly less liver steatosis and greater glucose tolerance and insulin sensitivity. obR mice also manifested lower food intake and greater energy expenditure than ob/ob mice, providing a mechanism underlying their metabolic improvement. Furthermore, obRs have sustained metabolic protection and restoration of fertility. Collectively, our studies show the importance of functional adipocytes in preventing metabolic abnormalities seen in leptin deficiency and point to the possibility of cell-based therapies for the treatment of leptin-deficient states.

Contribution of Adipose-Derived Factor D/Adipsin to Complement Alternative Pathway Activation: Lessons from Lipodystrophy.

Factor D (FD) is an essential component of the complement alternative pathway (AP). It is an attractive pharmaceutical target because it is an AP-specific protease circulating in blood. Most components of the complement activation pathways are produced by the liver, but FD is highly expressed by adipose tissue. Two critical questions are: 1) to what degree does adipose tissue contribute to circulating FD levels and 2) what quantity of FD is sufficient to maintain a functional AP? To address these issues, we studied a novel mouse strain with complete lipodystrophy (LD), the fld mouse with partial LD, an FD-deficient mouse, and samples from lipodystrophic patients. FD was undetectable in the serum of LD mice, which also showed minimal AP function. Reconstitution with purified FD, serum mixing experiments, and studies of partial LD mice all demonstrated that a low level of serum FD is sufficient for normal AP activity in the mouse system. This conclusion was further supported by experiments in which wild-type adipose precursors were transplanted into LD mice. Our results indicate that almost all FD in mouse serum is derived from adipose tissue. In contrast, FD levels were reduced ∼50% in the sera of patients with congenital generalized LD. Our studies further demonstrate that a relatively small amount of serum FD is sufficient to facilitate significant time-dependent AP activity in humans and in mice. Furthermore, this observation highlights the potential importance of obtaining nearly complete inhibition of FD in treating alternative complement activation in various autoimmune and inflammatory human diseases.

Glucocorticoid Receptor Signaling Is Not Required for In Vivo Adipogenesis.

Regulation of adipogenesis is of major interest given that adipose tissue expansion and dysfunction are central to metabolic syndrome. Glucocorticoids (GCs) are important for adipogenesis in vitro. However, establishing a role for GCs in adipogenesis in vivo has been difficult. GC receptor (GR)‒null mice die at birth, a time at which wild-type (WT) mice do not have fully developed white adipose depots. We conducted several studies aimed at defining the role of GC signaling in adipogenesis in vitro and in vivo. First, we showed that GR-null mouse embryonic fibroblasts (MEFs) have compromised ability to form adipocytes in vitro, a phenotype that can be partially rescued with a peroxisome proliferator-activated receptor γ agonist. Next, we demonstrated that MEFs are capable of forming de novo fat pads in mice despite the absence of GR or circulating GCs [by bilateral adrenalectomy (ADX)]. However, ADX and GR-null fat pads and their associated adipocyte areas were smaller than those in controls. Second, using adipocyte-specific luciferase reporter mice, we identified adipocytes in both WT and GR-null embryonic day (E)18 mouse embryos. Lastly, positive perilipin staining in WT and GR-null E18 embryos confirmed the presence of early white inguinal and brown adipocytes. Taken together, these results provide compelling evidence that GCs and GR augment but are not required for the development of functional adipose tissue in vivo.

Hepatic Glucocorticoid Receptor Plays a Greater Role Than Adipose GR in Metabolic Syndrome Despite Renal Compensation.

Exogenous glucocorticoid administration results in hyperglycemia, insulin resistance, hepatic dyslipidemia, and hypertension, a constellation of findings known as Cushing's syndrome. These effects are mediated by the glucocorticoid receptor (GR). Because GR activation in liver and adipose has been implicated in metabolic syndrome (MS), we wanted to determine the role of GR in these tissues in the development of MS. Because GR knockout (KO) mice (whole-body KO) exhibit perinatal lethality due to respiratory failure, we generated tissue-specific (liver or adipose) GRKO mice using cre-lox technology. Real-time PCR analysis of liver mRNA from dexamethasone-treated wildtype (WT) and liver GRKO mice indicated that hepatic GR regulates the expression of key genes involved in gluconeogenesis and glycogen metabolism. Interestingly, we have observed that liver-specific deletion of GR resulted in a significant increase in mRNA expression of key genes involved in gluconeogenesis and glycogen metabolism in kidney tissue, indicating a compensatory mechanism to maintain glucose homeostasis. We have also observed that GR plays an important role in regulating the mRNA expression of key genes involved in lipid metabolism. Liver GRKO mice demonstrated decreased fat mass and liver glycogen content compared with WT mice administered dexamethasone for 2 weeks. Adipose-specific deletion of GR did not alter glucose tolerance or insulin sensitivity of adipose GRKO mice compared with WT mice administrated dexamethasone. This indicates that liver GR might be more important in development of MS in dexamethasone-treated mice, whereas adipose GR plays a little role in these paradigms.